PROJECT SUMMARY/ABSTRACT The vital functions or respiration and digestion are carried out by organs that arise from an embryonic structure known as the gut tube. Despite its fundamental importance, morphogenesis of the gut tube has been remarkably understudied, particularly when considered alongside concurrent events in the neural tube, heart, limbs, and other tissues of the developing embryo. The purpose of this work is to study how this structure is formed through an integration of biophysical and molecular approaches, with the goal of revealing how progenitor cell movements in the endodermal epithelium are coordinated with differentiation to construct the foregut, midgut, and hindgut through molecular control of mechanical forces. The gut tube, once formed, is a simple epithelial cylinder, yet it arises through spatiotemporally distinct events that separately give rise to the foregut, midgut, and hindgut segments of the tube. These processes are poorly understood, and in particular, how the formation of these three segments are coordinated to form a single continuous tube remains largely unaddressed. Recently, we have turned to the chick embryo to study gut tube morphogenesis, employing live in vivo imaging, electroporation-based gene misexpression, and biomechanical/mathematical approaches to study formation of the hindgut. These studies revealed that much of what was previously assumed from fate mapping studies about gut tube formation may be incomplete, if not incorrect. The present application extends these approaches to study the biophysical role of collective cell movements in foregut morphogenesis, and to elucidate how cells of the presumptive midgut contribute cells to the forming foregut and hindgut. We will perform highly quantitative analyses of cell movements and physical forces in the presumptive foregut and midgut endoderm, and study the emergence of antero-posterior patterning in the context of cell movements along this axis, using single molecule fluorescent in situ detection of established marker genes. Understanding of this critical step in vertebrate development will provide valuable insight into the underlying causes of a broad range of gastrointestinal birth defects. Ultimately, the projects emanating from this work will have important implications for the development of cell-based therapies where directed the differentiation of progenitor cells toward distinct gastrointestinal and respiratory lineages is needed.